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Pyruvate Dehydrogenase (PDH) Activity Colorimetric Assay Kit

based on 4 citations in multiple journalsPyruvate Dehydrogenase (PDH) Activity Colorimetric Assay Kit44.1 4
Catalog #: K679

In stock

$475.00

Product Details

Cat # +Size K679-100
Size 100 assays
Detection Method Absorbance (450 nm)
Applications Measurement of pyruvate dehydrogenase activity in various tissues/cells
Analysis of cell signaling pathway
Features & Benefits • Simple, rapid & convenient
• can measure pyruvate dehydrogenase activity lower than 0.1 mU in a variety of samples
Kit Components • PDH Assay Buffer
• PDH Substrate (Lyophilized)
• PDH Developer (Lyophilized)
• NADH Standard (Lyophilized)
• PDH Positive Control
Storage Conditions -20°C
Shipping Conditions Gel Pack
USAGE For Research Use Only! Not For Use in Humans.

Details

Pyruvate Dehydrogenase (PDH) (EC 1.2.4.1) has a vital role in carbohydrate metabolism. It forms a well-characterized enzyme complex with dihydrolipoyl transacetylase (E2) and dihydrolipoyl dehydrogenase (E3). PDH converts pyruvate into acetyl-CoA in the presence of NAD and CoA, and links glycolysis to the citric acid cycle. PDH activity is inhibited by high intracellular ratios of ATP/ADP, NADH/NAD or Acetyl-CoA/CoA. In humans, PDH deficiency reduces mitochondrial function and is linked to neurodegenerative diseases. PDH deficiency is X-linked; it results in 2 forms of abnormality: a metabolic form (lactic acidosis) and a neurological form (seizure and/or neuropathological spasm). Recent studies show that PDH is a target of oncogene-induced senescence; activation of PDH enhances pyruvate utilization and increases respiration and redox stress. BioVision’s PDH assay kit provides a quick and easy way for monitoring PDH activity in various samples. In the assay, PDH converts pyruvate into an intermediate, which reduces the developer to a colored product with strong absorbance at 450 nm. The assay is simple, sensitive and can detect pyruvate dehydrogenase activity lower than 0.1 mU in a variety of samples.


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Can the supernatants from the lysed cellsbe frozen and stored until all the cells are ready to use so that all the samples can be run at the same time?
Although we always prefer fresh samples, the cell lysates can be stored at -80C before the assay. The lysates should be aliquoted if the assay will be repeated with the same samples to minimize freezing and thawing. Alternately, all cells can be stored at -80C and the lysates can be prepared on the day of the experiment.
Is it necessary to purify mitochondria to measure PDGH activity with this assay?
If you use total cell lysate, total PDH activity will be measured. Only if you fractionate the mitochondria and use that as sample, exclusively mitochondrial PDH activity will be measured.
On what species will this kit work?
Our kits are desgined and optimized with mammalian samples but this assay should measure PDH activity (pyruvate dehydrogenation is an evolutionarily conserved reaction) in any sample irrespective of species as long as there is PDH activity within the measurable range of the assay.
Can frozen samples be used with this assay?
Fresh samples are always preferred over frozen samples. However, frozen samples can also be used, provided, they were frozen right after isolation, were not freeze thawed multiple time (for which we recommend aliquoting the samples before freezing) and have been frozen for relatively short periods.
Is it possible to use a different wavelength than recommended for the final analysis?
It is always recommended to use the exact recommended wavelength for the most efficient results. However, most plate readers have flexibility in their band width of detection in increments of +/- 10 nm. Depending on this flexibility range, you can deviate from the recommended wavelengths within limits.
What is the exact volume of sample required for this assay?
There is no specific volume we can recommend for the amount any sample to be used since it is completely sample concentration and quality based. It is recommended to do a pilot expt with multiple sample volumes to determine the optimal volume which gives a reading within the linear range of the standard curve. Please refer to the citations for this product to see what other clients have used with similar sample types.
Are trial sizes of this kit available?
Unfortunately, we do not have trial sizes of this kit available. However,for customers based in the US or Canada, a 10% off list price introductory discount can be offered on the list price. For all other customers based out of this area our regional BioVision distributor should be contacted
What is the shelf life of this kit?
This kit is good for 12 months from the date of shipment in the unopened form when stored at the appropriate temperature and appropriate conditions. After opening and reconstitution, some of the components in this kit are good for 2 months at -20°C. Please refer to the datasheet for storage information and shelf life of each of the components.
Why are the standard curve values lower than those shown on the datasheet?
There are multiple factors which influence the signals like the incubation times, room temperature, handling etc. In general, to increase the value of the standards, the incubation time can be increased. As long as the standard curve is linear, it should be fine to use, since all the samples will also be measured under the same conditions on this curve.
Consitt, Leslie A. et al. (2016) Age-related impairments in skeletal muscle PDH phosphorylation and plasma lactate are indicative of metabolic inflexibility and the effects of exercise training. Am J Physiol Endocrinol Metab. 2016 Jul 1;311(1):E145-56.
Luz, Anthony L. et al. (2016) From the Cover: Arsenite Uncouples Mitochondrial Respiration and Induces a Warburg-like Effect in Caenorhabditis elegans. Toxicol Sci. 2016 Aug;152(2):349-62.
Sin et al., Acute Treatment of Resveratrol Alleviates Doxorubicin-Induced Myotoxicity in Aged Skeletal Muscle Through SIRT1-Dependent Mechanisms.  J Gerontol A Biol Sci Med Sci, Oct 2015; 10.1093/gerona/glv175.
Hagve et al., Skeletal muscle mitochondria exhibit decreased pyruvate oxidation capacity and increased ROS emission during surgery-induced acute insulin resistance. Am J Physiol Endocrinol Metab, Apr 2015; 308: E613 - E620.
For more citations of this product click here